Termites are social insects that subsist on sugars and other micronutrients obtained from nutritionally poor lignocellulose diets (Ohkuma M., (2006) Appl. Microbiol. Biotechnol. 61: 1-9; Scharf & Tartar (2008) Biofuels Bioprod. Birefin. 2: 540-552). Lignocellulose is a natural complex of the three biopolymers cellulose, hemicellulose and lignin. Cellulose is composed of long β-1,4-linked polymers of glucose that are held together in bundles by hemicellulose (Ljungdahl & Erickson (1985) Adv. Micro. Ecol. 8: 237-299; Lange J. P. (2007) Biofuels Bioprod. Bioref. 1: 39-48). Hemicellulose is composed of shorter β-1,4-linked polymers of mixed sugars such as mannose, xylose, galactose, rhamnose, arabinose, glucuronic acid, mannuronic acid, and galacturonic acid (Saha B. C., (2003) J. Indust. Microbiol. Biotechnol. 30: 279-291). Lignin is a 3-dimensional polymer of phenolic compounds that are linked to each other and to hemicellulose by ester bonds. Lignin is composed of three “mono-lignol” monomers (p-coumaryl alcohol, coniferyl alcohol, and sinapyl alcohol), which are combined in different ratios depending on the plant species. Another important characteristic of hemicellulose is its esterification with monomers and dimers of phenolic acid esters, which are identical to the mono-lignols that compose lignin (Saha B. C., (2003) J. Indust. Microbiol. Biotechnol. 30: 279-291; Crepin et al., (2004) Appl. Microbiol. Biotechnol. 63, 647-652; Benoit et al., (2008) Biotechnol. Letters 30, 387-396).
Termites digest lignocellulose with the assistance of endogenous and symbiont-produced digestive enzymes and co-factors (Breznak and Brune (1995) Appl. Env. Microbiol. 61: 2681-2687; Watanabe et al., (1998) Nature 394: 330-331; Ohkuma et al., (2006) Appl. Microbiol. Biotechnol. 61: 1-9; Scharf & Tartar (2008) Biofuels Bioprod. Birefin. 2: 540-552). Termite gut endosymbionts include a diversity of microorganisms that include protozoa, bacteria, spirochetes, fungi, and yeast, among others (Breznak and Brune (1995) Appl. Env. Microbiol. 61: 2681-2687; Warnecke et al., (2007) Nature 450, 560-565). The order Isoptera is divided into the higher and lower termites based mostly on symbiont composition. Lower termites, including Reticulitermes flavipes, possess cellulolytic protozoa in addition to a host of hydrogenic, methanogenic, and nitrogen fixing bacteria and spirochetes. Higher termites lack protozoa altogether, but instead possess cellulolytic bacteria. The roles of endosymbiotic fungi in higher and lower termites are not well defined; however, some higher termites cultivate fungus gardens in their nests that assist in lignocellulose digestion by producing cellulases, hemicellulases and lignases (Taprab et al., (2005) Appl. Env. Microbiol. 71: 7696-7704; Okhuma M., (2006) Appl Microbiol Biotechnol 61, 1-9).
Esterases are hydrolytic enzymes that cleave ester bonds in a diversity of biomolecules (Oakeshott et al. (2005) in Gilbert, L. I., Iatrou, K., Gill, S. S. (eds.) Comprehensive molecular insect science, Vol. 5, Elsevier-Pergamon, New York, pp. 309-382). Some insect esterases have very well defined biological functions, such as those involved in xenobiotic, lipid, acetylcholine, and juvenile hormone metabolism. However, many other insect esterases have largely undefined functions, yet are extremely efficient at metabolizing model substrates such as naphthyl and p-nitrophenyl esters. This latter category of esterases is referred to as the “general esterases”. Because of the highly esterified structure of lignin, it is possible that some general esterases may also contribute to lignin depolymerization in wood feeding insects such as termites.
Lignocellulose is a sustainable global resource with a great deal of relevance to renewable energy production. It is a naturally occurring complex of plant derived materials that includes the β-1,4-linked sugar polymers cellulose and hemicellulose, and the phenolic polymer lignin. In plants, lignocellulose provides key structural support for cell walls. Because it is plant-derived, lignocellulose is the most abundant and widespread bioenergy feedstock available on Earth. However, a major limitation in plant biomass utilization as a renewable energy source is the inefficiency of industrial lignocellulose depolymerization. This inefficiency increases energy inputs, reduces product yields, drives production costs higher, encourages political skepticism, and ultimately limits acceptance of cellulose-based renewable bioenergy. With respect to the problem of lignocellulose recalcitrance, it is germane that a number of invertebrate animals, and to some extent, their symbiotic gut fauna, have evolved specialized enzymes that cooperate in lignocellulose processing. In particular, endogenous lignocellulases encoded in marine and terrestrial invertebrate genomes can often confer high degrees of digestion capabilities to these organisms. When endogenous insect lignocellulases work synergistically with symbiont-derived enzymes, this can confer extremely high efficiency in lignocellulose processing. Termites (order Isoptera) are one of the most well recognized examples of an organism that subsists on lignocellulose; and thus, lignocellulase enzymes from termites and their gut symbionts have many potential bioenergy applications that warrant consideration.